Abstract
New protein folds have emerged throughout evolution, but it remains unclear how a protein fold can evolve while maintaining its function, particularly when fold changes require several sequential gene rearrangements. Here, we explored hypothetical evolutionary pathways linking different topological families of the DNA-methyltransferase superfamily. These pathways entail successive gene rearrangements through a series of intermediates, all of which should be sufficiently active to maintain the organism's fitness. By means of directed evolution, and starting from HaeIII methyltransferase (M.HaeIII), we selected all the required intermediates along these paths (a duplicated fused gene and duplicates partially truncated at their 5′ or 3′ coding regions) that maintained function in vivo. These intermediates led to new functional genes that resembled natural methyltransferases from three known classes or that belonged to a new class first seen in our evolution experiments and subsequently identified in natural genomes. Our findings show that new protein topologies can evolve gradually through multistep gene rearrangements and provide new insights regarding these processes.
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Acknowledgements
We thank the Minerva Foundation and the Israel Science Foundation for financial support, M. Babor for the software that generated the truncated PDB files and A. Levy for helpful comments on the manuscript. S.G.P. is the recipient of a Dewey D. Stone Postdoctoral Fellowship, L.R. is the recipient of a Feinberg Graduate School Fellowship and D.S.T. is the incumbent of the Elaine Blond Career Development Chair.
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Supplementary information
Supplementary Fig. 1
Fused Mtase dimers present in natural genomes. (PDF 143 kb)
Supplementary Fig. 2
Strategy used for the creation of truncated libraries using the ITCHY methodology. (PDF 11 kb)
Supplementary Fig. 3
In vivo methyltransferase activity of the disrupted N- and C-terminally truncated selected evolutionary intermediates. (PDF 77 kb)
Supplementary Fig. 4
Selection of Cluster I–derived circular permutants (PDF 39 kb)
Supplementary Fig. 5
In vivo methyltransferase activity of the disrupted Cluster I– and Cluster IV–derived selected circular permutants (PDF 48 kb)
Supplementary Fig. 6
Direct selection of circular permutants. (PDF 43 kb)
Supplementary Fig. 7
In vivo methyltransferase activity of the disrupted directly selected circular permutants. (PDF 62 kb)
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Peisajovich, S., Rockah, L. & Tawfik, D. Evolution of new protein topologies through multistep gene rearrangements. Nat Genet 38, 168–174 (2006). https://doi.org/10.1038/ng1717
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DOI: https://doi.org/10.1038/ng1717
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